Hyodeoxycholic Acid Research Articles

A Biomimetic Strategy for the Fabrication of Micro- and Nanodiamond Composite Films

This investigation is motivated by increasing interest in diamond and composite films for applications in biomedical and electronic devices. A biomimetic strategy is based on the use of commercial bile acids, such as ursodeoxycholic acid (UDCA) and hyodeoxycholic acid (HDCA). Composite films are developed using UDCA and HDCA as solubilizing agents for poly(ethyl methacrylate) (PEMA) in isopropanol and as dispersing agents for micro- and nanodiamonds. In this approach, the use of traditional toxic solvents for PEMA dissolution is avoided. The ability to obtain high concentrations of high molecular mass PEMA and disperse diamond particles in such solutions is a key factor for the development of a dip-coating method. The PEMA dissolution and diamond dispersion mechanisms are discussed. The composition and microstructure of the films can be varied by variation of the diamond particle size and concentration in the suspensions. The films can be obtained as singular layers of different compositions, multilayers of similar composition, or alternating layers of different compositions. The films combine corrosion protection property and biocompatibility of PEMA with advanced functional properties of diamonds.

Serum Metabolomic Profiling Reveals Biomarkers for Early Detection and Prognosis of Esophageal Squamous Cell Carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the most common aggressive malignancies worldwide, particularly in northern China. The absence of specific early symptoms and biomarkers leads to late-stage diagnosis, while early diagnosis and risk stratification are crucial for improving overall prognosis. We performed UPLC-MS/MS on 450 ESCC patients and 588 controls consisting of a discovery group and two validation groups to identify biomarkers for early detection and prognosis. Bioinformatics and clinical statistical methods were used for profiling metabolites and evaluating potential biomarkers. A total of 105 differential metabolites were identified as reliable biomarker candidates for ESCC with the same tendency in three cohorts, mainly including amino acids and fatty acyls. A predictive model of 15 metabolites [all-trans-13,14-dihydroretinol, (±)-myristylcarnitine, (2S,3S)-3-methylphenylalanine, 3-(pyrazol-1-yl)-L-alanine, carnitine C10:1, carnitine C10:1 isomer1, carnitine C14-OH, carnitine C16:2-OH, carnitine C9:1, formononetin, hyodeoxycholic acid, indole-3-carboxylic acid, PysoPE 20:3, PysoPE 20:3(2n isomer1), and resolvin E1] was developed by logistic regression after LASSO and random forest analysis. This model held high predictive accuracies on distinguishing ESCC from controls in the discovery and validation groups (accuracies > 89%). In addition, the levels of four downregulated metabolites [hyodeoxycholic acid, (2S,3S)-3-methylphenylalanine, carnitine C9:1, and indole-3-carboxylic acid] were significantly higher in early cancer than advanced cancer. Furthermore, three independent prognostic markers were identified by multivariate Cox regression analyses with and without clinical indicators: a high level of MG(20:4)isomer and low levels of 9,12-octadecadienoic acid and L-isoleucine correlated with an unfavorable prognosis; the risk score based on these three metabolites was able to stratify patients into low or high risk. Moreover, pathway analysis indicated that retinol metabolism and linoleic acid metabolism were prominent perturbed pathways in ESCC. In conclusion, metabolic profiling revealed that perturbed amino acids and lipid metabolism were crucial metabolic signatures of ESCC. Both panels of diagnostic and prognostic markers showed excellent predictive performances. Targeting retinol and linoleic acid metabolism pathways may be new promising mechanism-based therapeutic approaches. Thus, this study would provide novel insights for the early detection and risk stratification for the clinical management of ESCC and potentially improve the outcomes of ESCC.

Effect of Jinzhen granule on two coronaviruses: The novel SARS-CoV-2 and the HCoV-229E and the evidences for their mechanisms of action

BackgroundAlthough severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and human coronavirus 229E (HCoV-229E) pose a huge threat to human public health, no specific treatment is available. Jinzhen granule (JZ) is a traditional eight ingredients-Chinese medicine with prominent efficacy for treating viral-induced diseases. However, little is known about the antiviral effect and mechanism of JZ against SARS-CoV-2 and HCoV-229E. PurposeThis study aimed to reveal the antiviral effects of JZ against SARS-CoV-2 and HCoV-229E, and to further explore the underlying mechanisms regulating the host immune response. MethodsThe chromatographic separation of JZ was performed using a Shimadzu analytical high-performance liquid chromatograph with UV detection and Alltech ELSD 2000ES. We conducted cytopathic effect (CPE) and plaque reduction assays to evaluate the antiviral effect of JZ. A lethal human angiotensin converting enzyme 2 (hACE2) transgenic mouse model of SARS-CoV-2 was established to determine the protective effect of JZ on mortality and lung virus titers. Real-time quantitative PCR assays were used to analyze the expression of proinflammatory cytokines in vitro and in vivo. Western blotting was further performed to determine the activities on regulating the nuclear factor kappa B (NF-κB)/MAPK pathway. Finally, mitochondrial membrane potential assays, flow cytometry analysis and western blotting were used to assess the anti-apoptotic potency toward HCoV-229E infection. ResultsThe results showed that 13 chemical components were identified and five peaks were determined and quantitated (gallic acid 1.97 mg/g, baicalin 20.69 mg/g, glycyrrhizic acid 4.92 mg/g, hyodeoxycholic acid 4.86 mg/g, cholic acid 4.07 mg/g). We found that JZ exerted inhibitory potency against SARS-CoV-2 and HCoV-229E in vitro by using CPE and plaque reduction assays, and it was further found that JZ protected mice infected by SARS-CoV-2 from death and inhibited lung virus titers. JZ also significantly decreased the induction of inflammatory cytokines (IL-1α, IL-6, CCL-5 and MIP-1β), similar to the observed in vitro effect. Moreover, JZ suppressed the release of inflammatory cytokines in vitro and it decreased the protein expression of p-p38 MAPK, p-JNK, p-NF-κB p65 and p-IκBα induced by HCoV-229E and increased the expression of IκBα. Notably, JZ significantly protected HCoV-229E-infected Huh-7 cells from mitochondrial damage and decreased apoptotic cells. The activation of the mitochondria-mediated apoptotic pathway was inhibited by JZ, as shown by the reduced expression of cleaved caspase-9, caspase-3 and p-PARP. ConclusionsIn conclusion, JZ (gallic acid 1.97 mg/g, baicalin 20.69 mg/g, glycyrrhizic acid 4.92 mg/g, hyodeoxycholic acid 4.86 mg/g, cholic acid 4.07 mg/g) exhibited antiviral activities against SARS-CoV-2 and HCoV-229E by regulating the NF-κB/MAPK pathway and the mitochondria-mediated apoptotic pathway. These findings demonstrated the efficacy of JZ against CoVs and suggested JZ treatment as a novel clinical therapeutic strategy for COVID-19.

Taurolithocholic acid but not tauroursodeoxycholic acid rescues phagocytosis activity of bone marrow-derived macrophages under inflammatory stress.

Spinal cord injury (SCI) causes cell death and consequently the breakdown of axons and myelin. The accumulation of myelin debris at the lesion site induces inflammation and blocks axonal regeneration. Hematogenous macrophages contribute to the removal of myelin debris. In this study, we asked how the inflammatory state of macrophages affects their ability to phagocytose myelin. Bone marrow‐derived macrophages (BMDM) and Raw264.7 cells were stimulated with lipopolysaccharides (LPS) or interferon gamma (IFNγ), which induce inflammatory stress, and the endocytosis of myelin was examined. We found that activation of the TLR4‐NFκB pathway reduced myelin uptake by BMDM, while IFNγ‐Jak/STAT1 signaling did not. Since bile acids regulate lipid metabolism and in some cases reduce inflammation, our second objective was to investigate whether myelin clearance could be improved with taurolithocholic acid (TLCA), tauroursodeoxycholic acid or hyodeoxycholic acid. In BMDM only TLCA rescued myelin phagocytosis, when this activity was suppressed by LPS. Inhibition of protein kinase A blocked the effect of TLCA, while an agonist of the farnesoid X receptor did not rescue phagocytosis, implicating TGR5‐PKA signaling in the effect of TLCA. To shed light on the mechanism, we measured whether TLCA affected the expression of CD36, triggering receptor on myeloid cells‐2 (TREM2), and Gas6, which are known to be involved in phagocytosis and affected by inflammatory stimuli. Concomitant with an increase in expression of tumour necrosis factor alpha, LPS reduced expression of TREM2 and Gas6 in BMDM, and TLCA significantly diminished this downregulation. These findings suggest that activation of bile acid receptors may be used to improve myelin clearance in neuropathologies.

Hyodeoxycholic acid rescues lipopolysaccharide-induced neuroinflammation via regulating TGR5/AKT/NF-κB pathway

Hyodeoxycholic acid (HDCA) is a natural secondary bile acid with enormous pharmacological effects. It can act as a takeda G-coupled protein receptor 5 (TGR5) agonist, activation of TGR5 is found to alleviate neuroinflammation by suppressing serine/threonine kinase (AKT)/ nuclear factor-κB (NF-κB) signaling pathway. In this study, the anti-microglial inflammatory effect of HDCA was evaluated in lipopolysaccharide (LPS)-induced BV2 microglial cells and mice (n=12/group). One-way ANOVA followed by Tukey’s post hoc analysis was conducted. p <.05 is statistically significant. HDCA was shown to inhibit the over-activation of microglia both in vitro and in vivo , evidenced by down-regulated mRNA expression of inflammatory cytokines such as interleukin 1β, decreased protein expression of inducible nitric oxide synthase and cyclooxygenase 2. Moreover, in LPS-induced microglial cells, HDCA was found to counteract the suppression of TGR5 and the increased phosphorylation of AKT, NF-κB and NF-κB inhibitor factor, as well as the nuclear translocation of NF-κB, however, the effect of HDCA onTGR5, AKT and NF-κB was abolished by the TGR5 inhibitor - triamterene. HDCA suppresses LPS-induced neuroinflammation through regulating TGR5/AKT/NF-κB signaling pathway. The novel findings suggest the therapeutic potential of HDCA in central nervous system disorders with prominent neuroinflammation.

Analysis of eight bile acids in urine of gastric cancer patients based on covalent organic framework enrichment coupled with liquid chromatography-tandem mass spectrometry

Gastric carcinoma is one of the most common and deadly forms of cancer. Early detection is critical for successful treatment of gastric cancer, and examination of BAs in urine may provide a critical diagnostic tool for identifying gastric cancer at stages when it can still be cured. Bile acids (BAs) are a crucial toxic factor correlated with the injury of gastric mucosa and as such, quantifying the amount of BA in patient's urine could provide a new means to quickly and non-invasively identify the presence of gastric cancer in the early stages. Here, a covalent organic framework (COF) material synthesized on the basis of 1,3,5-tris(4-nitrophenyl)benzene (TAPB) and pyromellitic dianhydride (PMDA) was used as stationary phase for SPE column that was coupled to LC-MS/MS for quantitative analysis of eight BAs in human urine, including cholic acid (CA), deoxycholic acid (DCA), glycochenodeoxycholic acid (GCDCA), glycocholic acid (GCA), taurochenodeoxycholic acid (TCDCA), lithocholic acid (LCA), hyodeoxycholic acid (HDCA), and chenodeoxycholic acid (CDCA). The enrichment effect of synthesized COF material was better than commercial SPE and HLB column. The sensitivity can increase 9.37- to 54.30- fold (calculated by the ratio of peak area between before and after enrichment). The probable mechanism is due to the great porosity and the similar polarity with BAs of the COF material. By compared with previous literatures, our method had the minimum limit of detection, which achieved 46.40, 25.75, 47.40, 47.37, 30.42, and 33.92 pg /mL, respectively, for GCA, GCDCA, CA, CDCA, HDCA and DCA after enrichment. These eight BAs also accomplished excellent linearity from 0.34 to 10,000 ng/mL. This material was successfully applied in the measurements of these six BAs in human urine from 76 gastric cancer patients and 32 healthy people. Compared to healthy people, levels of CA, CDCA, DCA, and HDCA were significantly elevated and levels of GCDCA were depressed, respectively, in gastric cancer patients. Our work suggests that these acids may act as potential biomarkers for gastric cancer and our framework provides a method for “non-invasive” diagnosis of gastric cancer.

Plasma Bile Acid Profile in Patients with and without Type 2 Diabetes.

A paucity of information currently exists on plasma bile acid (BA) profiles in patients with and without type 2 diabetes mellitus (T2DM). We assayed 14 plasma BA species in 224 patients with T2DM and in 102 nondiabetic individuals with metabolic syndrome. Plasma BA levels were measured with ultra-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) technique. Multivariable linear regression analyses were undertaken to assess associations between measured plasma BA species and T2DM status after adjustment for confounding factors. The presence of T2DM was significantly associated with higher plasma concentrations of both primary BAs (adjusted-standardized β coefficient: 0.279, p = 0.005) and secondary BAs (standardized β coefficient: 0.508, p < 0.001) after adjustment for age, sex, adiposity measures, serum alanine aminotransferase and use of statins or metformin. More specifically, the presence of T2DM was significantly associated with higher levels of plasma taurochenodeoxycholic acid, taurodeoxycholic acid, glycochenodeoxycholic acid, hyodeoxycholic acid, glycodeoxycholic acid, glycolithocholic acid, deoxycholic acid, taurochenodeoxycholic acid, taurodeoxycholic acid, glycochenodeoxycholic acid and glycodeoxycholic acid (adjusted-standardized β coefficients ranging from 0.315 to 0.600; p < 0.01 or less), as well as with lower plasma levels of cholic acid (adjusted-standardized β coefficient: −0.250, p = 0.013) and taurocholic acid (adjusted-standardized β coefficient: −0.309, p = 0.001). This study shows that there are marked differences in plasma BA profiles between patients with and without T2DM. Further research will be needed to better understand how these differences in plasma BA profiles may interplay with the pathophysiology of T2DM.

Detection Bioactive Metabolites of Fructobacillus fructosus Strain HI-1 Isolated from Honey Bee's Digestive Tract Against Paenibacillus larvae.

American foulbrood is a devastating disease of honey bee, causing economic loss in the beekeeping industry. The disease mainly causes reduction in honeybee populations which negatively affect the honey bee's major role as natural pollinators of significant crops and wildflowers. Thus, it is crucial to develop safe efficient strategies to control the disease and to improve bee colony health. Using lactic acid bacteria (LAB) as an alternative to chemical treatments is a promising novel technique for tackling honeybee diseases and improving their immunity. The endogenous LAB isolates were recovered from honeybee gut samples collected from different apiaries in two Egyptian governorates and screened for antagonistic activities against Paenibacillus larvae (pathogen of AFB disease). The results showed that 53.3% of tested LAB isolates (n = 120) exhibited antagonistic activities against P. larvae. The minimum inhibitory concentration and minimum bactericidal concentration of the most potent LAB isolate (with an inhibition zone of 44 mm) were 100 and 125 µL/mL, respectively. 16S rRNA sequencing identified the most potent isolate as Fructobacillus fructosus HI-1. The bioactive metabolites of F. fructosus were extracted with ethyl acetate and fractionated on thin-layer chromatography (TLC); also, bioactive fractions were detected. Heptyl 2-methylbutyrate, di-isobutyl phthalate, D-turanose, heptakis (trimethylsilyl), di-isooctyl phthalate, and hyodeoxycholic acid compounds were identified in the bioactive fractions. The result explores the promising administration of probiotic metabolites to control honeybee AFB disease, as a natural tool to substitute antibiotics and chemicals in disease-controlling strategies.

Exposure to High Aerial Ammonia Causes Hindgut Dysbiotic Microbiota and Alterations of Microbiota-Derived Metabolites in Growing Pigs.

Ammonia, an atmospheric pollutant in the air, jeopardizes immune function, and perturbs metabolism, especially lipid metabolism, in human and animals. The roles of intestinal microbiota and its metabolites in maintaining or regulating immune function and metabolism are irreplaceable. Therefore, this study aimed to investigate how aerial ammonia exposure influences hindgut microbiota and its metabolites in a pig model. Twelve growing pigs were treated with or without aerial ammonia (35 mg/m3) for 25 days, and then microbial diversity and microbiota-derived metabolites were measured. The results demonstrated a decreasing trend in leptin (p = 0.0898) and reduced high-density lipoprotein cholesterol (HDL-C, p = 0.0006) in serum after ammonia exposure. Besides, an upward trend in hyocholic acid (HCA), lithocholic acid (LCA), hyodeoxycholic acid (HDCA) (p < 0.1); a downward trend in tauro-deoxycholic acid (TDCA, p < 0.1); and a reduced tauro-HDCA (THDCA, p < 0.05) level were found in the serum bile acid (BA) profiles after ammonia exposure. Ammonia exposure notably raised microbial alpha-diversity with higher Sobs, Shannon, or ACE index in the cecum or colon and the Chao index in the cecum (p < 0.05) and clearly exhibited a distinct microbial cluster in hindgut indicated by principal coordinate analysis (p < 0.01), indicating that ammonia exposure induced alterations of microbial community structure and composition in the hindgut. Further analysis displayed that ammonia exposure increased the number of potentially harmful bacteria, such as Negativibacillus, Alloprevotella, or Lachnospira, and decreased the number of beneficial bacteria, such as Akkermansia or Clostridium_sensu_stricto_1, in the hindgut (FDR < 0.05). Analysis of microbiota-derived metabolites in the hindgut showed that ammonia exposure increased acetate and decreased isobutyrate or isovalerate in the cecum or colon, respectively (p < 0.05). Unlike the alteration of serum BA profiles, cecal BA data showed that high ammonia exposure had a downward trend in cholic acid (CA), HCA, and LCA (p < 0.1); a downward trend in deoxycholic acid (DCA) and HDCA (p < 0.05); and an upward trend in glycol-chenodeoxycholic acid (GCDCA, p < 0.05). Mantel test and correlation analysis revealed associations between microbiota-derived metabolites and ammonia exposure-responsive cecal bacteria. Collectively, the findings illustrated that high ammonia exposure induced the dysbiotic microbiota in the hindgut, thereby affecting the production of microbiota-derived short-chain fatty acids and BAs, which play a pivotal role in the modulation of host systematic metabolism.

The Nonsteroidal Anti-Inflammatory Drug Ketorolac Alters the Small Intestinal Microbiota and Bile Acids Without Inducing Intestinal Damage or Delaying Peristalsis in the Rat.

Background: Nonsteroidal anti-inflammatory drugs (NSAIDs) induce significant damage to the small intestine, which is accompanied by changes in intestinal bacteria (dysbiosis) and bile acids. However, it is still a question of debate whether besides mucosal inflammation also other factors, such as direct antibacterial effects or delayed peristalsis, contribute to NSAID-induced dysbiosis. Here we aimed to assess whether ketorolac, an NSAID lacking direct effects on gut bacteria, has any significant impact on intestinal microbiota and bile acids in the absence of mucosal inflammation. We also addressed the possibility that ketorolac-induced bacterial and bile acid alterations are due to a delay in gastrointestinal (GI) transit. Methods: Vehicle or ketorolac (1, 3 and 10 mg/kg) were given to rats by oral gavage once daily for four weeks, and the severity of mucosal inflammation was evaluated macroscopically, histologically, and by measuring the levels of inflammatory proteins and claudin-1 in the distal jejunal tissue. The luminal amount of bile acids was measured by liquid chromatography-tandem mass spectrometry, whereas the composition of microbiota by sequencing of bacterial 16S rRNA. GI transit was assessed by the charcoal meal method. Results: Ketorolac up to 3 mg/kg did not cause any signs of mucosal damage to the small intestine. However, 3 mg/kg of ketorolac induced dysbiosis, which was characterized by a loss of families belonging to Firmicutes (Paenibacillaceae, Clostridiales Family XIII, Christensenellaceae) and bloom of Enterobacteriaceae. Ketorolac also changed the composition of small intestinal bile by decreasing the concentration of conjugated bile acids and by increasing the amount of hyodeoxycholic acid (HDCA). The level of conjugated bile acids correlated negatively with the abundance of Erysipelotrichaceae, Ruminococcaceae, Clostridiaceae 1, Muribaculaceae, Bacteroidaceae, Burkholderiaceae and Bifidobacteriaceae. Ketorolac, under the present experimental conditions, did not change the GI transit. Conclusion: This is the first demonstration that low-dose ketorolac disturbed the delicate balance between small intestinal bacteria and bile acids, despite having no significant effect on intestinal mucosal integrity and peristalsis. Other, yet unidentified, factors may contribute to ketorolac-induced dysbiosis and bile dysmetabolism.

Mechanism of Qingkailing on influenza based on network pharmacology and molecular docking

Objective To explore the potential mechanism of Qingkailing (QKL) on influenza, and to provide a theoretical basis for the clinical application of QKL. Methods TCMSP, TCMID, and PubChem databases were used to search for the active ingredients and action targets of QKL. GeneCards database was used to search for the targets of influenza. The intersection method was used to obtain the targets related to the therapeutic effects of QKL. Cytoscape software was applied for the construction of active compounds-targets network map. Protein-protein interaction network was constructed by STRING database. Gene ontology functional enrichment analysis and KEGG pathway enrichment analysis were conducted by Bioconductor database and R software. Auto Dock Tools were used for molecular docking. Results Total 90 potential active components were identified from QKL with the corresponding 225 targets. PPI network analysis showed that there were 34 key targets intervening influenza by QKL. GO and KEGG showed that the mechanism of QKL intervention on influenza was related to anti-inflammatory and antiviral. The results of molecular docking showed that cholic acid, hyodeoxycholic acid and baicalin had affinity with RELA and JUN. Conclusion The active ingredients of QKL target on JUN, RELA, MAPK1, IL6 and AKT1 to regulate multiple signal pathways, and play an intervention role in influenza.

Untargeted metabolomics for uncovering plasma biological markers of wet age-related macular degeneration.

Wet age-related macular degeneration (wAMD) causes central vision loss and represents a major health problem in elderly people. Here we have used untargeted metabolomics using UHPLC-MS to profile plasma from 127 patients with wAMD (67 choroidal neovascularization (CNV) and 60 polypoidal choroidal vasculopathy (PCV)) and 50 controls. A total of 545 biochemicals were detected. Among them, 17 metabolites presented difference between patients with wAMD and controls. Most of them were oxidized lipids (N=6, 35.29%). Comparing to controls, 28 and 18 differential metabolites were identified in patients with CNV and PCV, respectively. Two metabolites, hyodeoxycholic acid and L-tryptophanamide, were differently distributed between PCV and CNV. We first investigated the genetic association with metabolites in wet AMD (CFH rs800292 and HTRA1 rs10490924). We identified six differential metabolites between the GG and AA genotypes of CFH rs800292, five differential metabolites between the GG and AA genotypes of HTRA1 rs10490924, and four differential metabolites between the GG and GA genotypes of rs10490924. We selected four metabolites (cyclamic acid, hyodeoxycholic acid, L-tryptophanamide and O-phosphorylethanolamine) for in vitro experiments. Among them, cyclamic acid reduced the activity, inhibited the proliferation, increased the apoptosis and necrosis in human retinal pigment epithelial cells (HRPECs). L-tryptophanamide affected the proliferation, apoptosis and necrosis in HRPECs, and promoted the tube formation and migration in primary human retinal endothelial cells (HRECs). Hyodeoxycholic acid and O-phosphorylethanolamine inhibited the tube formation and migration in HRECs. The results suggested that differential metabolites have certain effects on wAMD pathogenesis-related HRPECs and HRECs.

Reduced Bile Acid Signaling Contributes to Hypertension Development in Spontaneously Hypertensive Stroke‐Prone Rats

Exciting recent findings have identified that gut microbiota plays an essential role in blood pressure (BP) regulation. Studies from our lab and others have shown a causal role of gut dysbiosis in hypertension development in the spontaneously hypertensive stroke-prone (SHRSP) rat and several other animal models. One key mechanism by which the microbiota influences the host is through the generation/modification of metabolites. Among these metabolites, bile acids (BAs), are derived from the liver and modified by gut bacteria. Many BA species cross the gut epithelial barrier into the host circulation and influence inflammation, metabolism, and vascular function through the binding and activation of BA receptors, including TGR5. We hypothesized that disrupted BA signaling, as a result of gut dysbiosis, contributes to the development of hypertension. We observed a significant reduction of several BAs in systemic plasma of SHRSP as compared to WKY, including the TGR5 agonists, cholic acid (CA), hyocholic acid (HCA), hyodeoxycholic acid (HDCA), and tauro-lithocholic acid (TLCA) (p<.05). We next supplemented WKY and SHRSP rats with 0.5% CA in the diet and measured systolic blood pressure (SBP) bi-weekly. After 15 weeks of treatment, systemic plasma was collected for BA analysis and endothelium-dependent dilation was assessed in the isolated aorta. We observed that CA supplementation significantly increased CA, HCA, HDCA, and TLCA in systemic plasma of SHRSP (p<.0001). CA supplementation also significantly reduced SBP of SHRSPs, with an average decrease of 23 mmHg over the final 6 weeks of treatment (p<.05) compared to SHRSP controls. Chronic CA treatment also significantly improved vasodilatory response to increasing doses of acetylcholine (ACh) in the isolated aorta of SHRSPs (p<.05). Next, we determined if BAs had their beneficial effects through activation of the BA receptor TGR5. Beginning at 6 weeks of age, SHRSPs were randomized into vehicle control or oleanolic acid (OA, 10 mg/kg IP daily), a selective TGR5 agonist, treatment groups. OA significantly decreased SBP by an average of 15 mmHg over the final 6 weeks of the treatment, compared to controls (p<.05). Furthermore, 15 weeks of OA treatment significantly improved vasodilatory response to increasing doses of ACh in isolated mesenteric arteries of SHRSPs (p<.05). Because systemic and neuronal inflammation also contributes to the development of hypertension, we examined the effects of TGR5 activation by OA treatment on inflammation in the gut and brain of SHRSPs. Using flow cytometry, we determined that OA treatment increased the percentage of anti-inflammatory Tregs and decreased pro-inflammatory T helper 17 cells in both cecum and brain of SHRSP (p<.05). OA treatment also significantly reduced interferon-γ+ T cells in the cecum of SHRSP. These data suggest an anti-inflammatory role of TGR5 signaling in the gut and brain. We conclude that reduced BA-mediated TGR5 signaling contributes to the development of hypertension in SHRSPs, and that activating TGR5 may be a potential therapeutic approach to attenuate vascular endothelial dysfunction and associated hypertension.

Synthesis and Biological Activity of New Brassinosteroid Analogs of Type 24-Nor-5β-Cholane and 23-Benzoate Function in the Side Chain.

Brassinosteroids are polyhydroxysteroids that are involved in different plants’ biological functions, such as growth, development and resistance to biotic and external stresses. Because of its low abundance in plants, much effort has been dedicated to the synthesis and characterization of brassinosteroids analogs. Herein, we report the synthesis of brassinosteroid 24-nor-5β-cholane type analogs with 23-benzoate function and 22,23-benzoate groups. The synthesis was accomplished with high reaction yields in a four-step synthesis route and using hyodeoxycholic acid as starting material. All synthesized analogs were tested using the rice lamina inclination test to assess their growth-promoting activity and compare it with those obtained for brassinolide, which was used as a positive control. The results indicate that the diasteroisomeric mixture of monobenzoylated derivatives exhibit the highest activity at the lowest tested concentrations (1 × 10−8 and 1 × 10−7 M), being even more active than brassinolide. Therefore, a simple synthetic procedure with high reaction yields that use a very accessible starting material provides brassinosteroid synthetic analogs with promising effects on plant growth. This exploratory study suggests that brassinosteroid analogs with similar chemical structures could be a good alternative to natural brassinosteroids.

Hyocholic acid and glycemic regulation: comments on ‘Hyocholic acid species improve glucose homeostasis through a distinct TGR5 and FXR signaling mechanism’

Hyocholic acid species (HCA, hyodeoxycholic acid, and their glycine and taurine conjugated forms) comprise 80% of the composition of pig bile (Haslewood, 1956). An interesting fact about pigs is that they do not get diabetes even though they eat almost anything and in abundant amounts, a diabetes-promoting diet. The first use of pig bile for treatment of 'xiao-ke', a condition known today as diabetes, was recorded ∼400 years ago by the Chinese medical practitioners in the Compendium of Materia Medica (Li, 1573‒1593). Recently, we found HCA species as novel biomarkers for metabolic diseases (Zheng et al., 2021b) and also identified the role of HCA species in the prevention of diabetes as well as their mechanism of action (Zheng et al., 2021a). Although bile acids (BAs) are mostly associated with their aid in food digestion, they have also been shown to act as signaling molecules by binding to two particular receptors, farnesoid X receptor (FXR) and the G-protein-coupled receptor, TGR5. Experiments were thus directed to the effect of HCA binding to these two BA receptors on glycemic regulation in both in vivo and in vitro models.

Isomaltulose Exhibits Prebiotic Activity, and Modulates Gut Microbiota, the Production of Short Chain Fatty Acids, and Secondary Bile Acids in Rats.

In vitro experiments have indicated prebiotic activity of isomaltulose, which stimulates the growth of probiotics and the production of short chain fatty acids (SCFAs). However, the absence of in vivo trials undermines these results. This study aims to investigate the effect of isomaltulose on composition and functionality of gut microbiota in rats. Twelve Sprague–Dawley rats were divided into two groups: the IsoMTL group was given free access to water containing 10% isomaltulose (w/w), and the control group was treated with normal water for five weeks. Moreover, 16S rRNA sequencing showed that ingestion of isomaltulose increased the abundances of beneficial microbiota, such as Faecalibacterium and Phascolarctobacterium, and decreased levels of pathogens, including Shuttleworthia. Bacterial functional prediction showed that isomaltulose affected gut microbial functionalities, including secondary bile acid biosynthesis. Targeted metabolomics demonstrated that isomaltulose supplementation enhanced cholic acid concentration, and reduced levels of lithocholic acid, deoxycholic acid, dehydrocholic acid, and hyodeoxycholic acid. Moreover, the concentrations of propionate and butyrate were elevated in the rats administered with isomaltulose. This work suggests that isomaltulose modulates gut microbiota and the production of SCFAs and secondary bile acids in rats, which provides a scientific basis on the use of isomaltulose as a prebiotic.

2,3,5,4'-Tetrahydroxystibane-2-O-β-D-glucoside induces liver injury by disrupting bile acid homeostasis and phospholipids efflux

Polygonum multiflorum is a traditional Chinese herbal medicine and has many biological activities such as hair-blacking, anti-atherosclerosis, anti-inflammatory and anti-aging. However, the liver injury induced by P. multiflorum has aroused wide attention in recent years. 2,3,5,4'-tetrahydroxystibane-2-O-β-D-glucoside(TSG) is a main component of P. multiflorum, but the role of TSG in inducing liver injury is unclear. The aim of present study was to evaluate TSG's potential liver injury and effects on bile acid homeostasis and phospholipids efflux. C57 BL/6 J mice received intraperitoneal administration of 400 mg·kg~(-1) of TSG daily for 15 days, and then biochemical indexes of liver injury and changes of phospholipid content were detected. The changes of bile acid compositions were detected by LC-MS/MS. The results showed TSG 400 mg·kg~(-1) significantly increased the content of serum total bile acid(TBA) and alkaline phosphatase(ALP). Elevated free bile acid levels were observed in TSG-treated groups, including β-muricholic acid(β-MCA), ursodeoxycholic acid(UDCA), hyodeoxycholic acid(HDCA), chenodeoxycholic acid(CDCA), deoxcholic acid(DCA) in serum and β-MCA, CDCA in liver. TSG inhibited the protein expression of farnesoid X receptor(FXR) and down stream bile salt export pump(BSEP), which may result in the accumulation of bile acid. TSG also inhibited the expression of 25-hydroxycholesterol-7 alpha-hydroxylase(CYP7 B1), which may disturb the alternative pathway for bile acid synthesis. In addition, intraperitoneal injection of TSG 400 mg·kg~(-1) significantly decreased the content of phospholipids in bile. The research showed that TSG significantly inhibited the expression of multidrug resistance protein 2(MDR2) and destroyed the regular distribution of MDR2 on the bile duct membrane of liver. In vitro results showed that the IC_(50) of TSG on HepG2 cells was about 1 500 μmol·L~(-1) and TSG at 500 μmol·L~(-1)(for 24 h) could destroy the distribution of MDR2 on the bile duct membrane of liver. In conclusion, TSG induced liver injury by disrupting bile acid homeostasis and phospholipids efflux.

Synthesis and biological activity of cyclopropyl Δ7-dafachronic acids as DAF-12 receptor ligands.

The four cyclopropyl stereoisomers of Δ7-dafachronic acids were prepared from the bile acid hyodeoxycholic acid and employed as chemical tools to exploit the importance of the orientation and spatial disposition of the carboxyl tail and the C25-methyl group for the binding at the DAF-12 receptor. The synthesis route was based on (a) Walden inversion and stereoselective PtO2-hydrogenation to convert the L-shaped 5β-cholanoid scaffold into the planar 5α-sterol intermediate; (b) two-carbon homologation of the side chain by Wittig and cyclopropanation reaction; and (c) formation of the 3-keto group and Δ7 double bond. The synthesized isomers were isolated and tested for their activity as DAF-12 ligands by AlphaScreen assays. Results showed a significant loss of potency and efficacy for all the four stereoisomers when compared to the parent endogenous ligand. Computational analysis has evidenced the configurational and conformational arrangement of both the carboxylic and the C25-methyl group of dafachronic acids as key structural determinants for DAF-12 binding and activation.

Apple polyphenols extract alleviated dextran sulfate sodium-induced ulcerative colitis in C57BL/6 male mice by restoring bile acid metabolism disorder and gut microbiota dysbiosis.

To investigate and compare the preventive effects of apple polyphenols extract (APE) with phloretin on dextran sulfate sodium (DSS)-induced ulcerative colitis (UC), 60 male mice were treated with 125 or 500 mg/(kg bw d) APE or 100 mg/(kg bw d) phloretin, the single-ingredient of APE, for continuous 3 weeks by intragastric administration, meanwhile, mice were provided with 3% DSS dissolved in drinking water to induce UC during the third week. Both APE and phloretin significantly ameliorated DSS-induced UC by inhibiting body weight loss, preventing colon shortening and mucosa damage. Except the same mechanisms of the inhibited activation of NF-κB signaling, decreased hyodeoxycholic acid level and increased abundance of Verrucomicrobia at phylum and Bacteroides and Akkermansia at genus, APE increased β-muricholic acid level and decreased Bacterodetes abundance, while phloretin decreased Firmicutes abundance. Furthermore, APE treatment showed much lower disease activity index score, less body weight loss and lighter spleen than phloretin. Thus, our study supported the potentiality of APE as a promising dietary intervention for the prevention of experimental UC.

Epimeric Mixtures of Brassinosteroid Analogs: Synthesis, Plant Growth, and Germination Effects in Tomato (Lycopersicum esculentum Mill.)

Brassinosteroids (BRs) play an important role in the growth and development of plants. Herein, we describe the synthesis of epimeric mixtures of BR analogs with 24-norcholane type side chains, S/R configuration at C22 and A/B ring cis-type fusion. All epimeric mixtures were synthetized from hyodeoxycholic acid. The biological activity of mixtures was evaluated by using rice lamina inclination test and germination of tomato (Lycopersicum esculentum) seeds. The results show that these epimeric mixtures exhibit similar bioactivity to brassinolide in both bioassays. Thus, our results corroborate that the A/B junction has almost no effect on bioactivity and open the possibility of using epimeric mixtures instead of pure compounds. In this approach, the synthesized BR analogs maintain a good level of bioactivity, whereas the synthesis is shorter, cheaper and with higher yields. All these factors make this alternative very interesting for potential application.